System And Method For Plasma Purification Prior To Mononuclear Cell Collection

ABSTRACT

A method of collecting mononuclear cells includes separating whole blood into plasma and cellular components, purifying the plasma through a plasma adsorption column to create purified plasma, combining the cellular components with the purified plasma to form a first mixture, and separating the first mixture into mononuclear cells and at least one component. Alternatively, whole blood may be flowed through an adsorption column to create purified whole blood, with the purified whole blood then being separated into mononuclear cells and at least one component.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Patent App. No.62/277,198 filed Jan. 11, 2016, which is expressly incorporated hereinby reference in its entirety.

FIELD OF THE DISCLOSURE

The present disclosure is directed to fluid treatment systems andmethods. More particularly, the present disclosure relates to systemsand methods for separating blood into its constituents and subsequentlytreating and/or collecting the constituents.

BACKGROUND

A variety of available blood processing systems allows for thecollection and processing of particular blood components, rather thanwhole blood, from donors or patients. In the case of a blood donor,whole blood is drawn from the donor, a desired blood constituentseparated and collected, and the remaining blood components returned tothe donor. By removing only particular constituents rather than wholeblood, it takes the donor's body a shorter time period to recover tonormal blood levels, thereby increasing the frequency with which thedonor may donate blood. It is beneficial to increase in this manner theoverall supply of blood constituents made available for health care,such as red blood cells (RBCs), leukocytes, mononuclear cells (MNCs),plasma, and/or platelets, etc. In the case of a patient, whole blood issimilarly drawn from the patient, a particular blood constituent firstseparated and then collected and/or treated, and the remaining bloodcomponents returned to the patient. The collected and/or treated bloodconstituent may be saved for future use, returned to the patient, and/ordiscarded and replaced with a suitable replacement.

The separation of blood components from whole blood typically takesplace prior to the collection or treatment of the separated bloodcomponent and may be achieved through a spinning membrane orcentrifugation, in which whole blood is passed through a centrifuge ormembrane after it is withdrawn from the patient. To avoid contaminationand possible infection of the patient, the blood is preferably containedwithin a sealed, sterile fluid flow system during the entire separationprocess. Typical blood processing systems thus may include a permanent,reusable hardware assembly containing the hardware (drive system, pumps,valve actuators, programmable controller, and the like) that pumps theblood, and a disposable, sealed and sterile fluid circuit that ismounted in cooperation on the hardware. In the case of separation viacentrifugation, the hardware assembly includes a centrifuge that mayengage and spin a separation chamber of the disposable fluid circuitduring a blood separation step. The blood, however, may make actualcontact only with the fluid circuit, which assembly may be used onlyonce and then discarded. In the case of separation via a spinningmembrane, a disposable single-use spinning membrane may be used incooperation with the hardware assembly and disposable fluid circuit.

In the case of separation via centrifugation, as the whole blood is spunby the centrifuge, the heavier (greater specific gravity) components,such as red blood cells, move radially outwardly away from the center ofrotation toward the outer or “high-G”wall of the separation chamber ofthe fluid circuit. The lighter (lower specific gravity) components, suchas plasma, migrate toward the inner or “low-G” wall of the separationchamber. Various ones of these components can be selectively removedfrom the whole blood by forming appropriately located channeling sealsand outlet ports in the separation chamber of the fluid circuit.

In the case of separation via a spinning membrane, whole blood may bespun within a disposable spinning membrane, rather than within aseparation chamber of a fluid circuit. Larger molecules, such as redblood cells, may be retained within one side of the membrane, while thesmaller molecules, such as plasma, may escape through the pores of themembrane to the other side of the membrane. Various ones of thesecomponents can be selectively removed from the whole blood by formingappropriately located outlet ports in the housing of the membranecolumn. Various types of membranes with different pore sizes may beused, depending on the components to be separated.

In the case of MNC collection, which includes the collection oflymphocytes, monocytes, and/or stem cells, MNCs can be removed from thewhole blood of a patient, collected, and/or subjected to varioustherapies. Collected and treated MNCs may then be returned to thepatient for the treatment of various blood diseases by, e.g.,eliminating immunogenicity in cells, inactivating or killing selectedcells, inactivating viruses or bacteria, reconstituting the immunesystem, and/or activating desirable immune responses. MNC treatments areused for blood or solid organ/tissue cancers, photopheresis treatments,autologous and allogeneic stem cell transplants, donor lymphocyteinfusions, research collections, etc.

SUMMARY

According to an exemplary embodiment, the present disclosure is directedto a method of collecting mononuclear cells, comprising separating wholeblood into plasma and cellular components, combining the cellularcomponents with plasma replacement fluid to form a first mixture, andseparating the first mixture into mononuclear cells and at least onecomponent.

According to an exemplary embodiment, the present disclosure is directedto a method of collecting mononuclear cells, comprising separating wholeblood into plasma and cellular components, purifying the plasma througha plasma adsorption column to create purified plasma, combining thecellular components with the purified plasma to form a first mixture,and separating the first mixture into mononuclear cells and at least onecomponent.

According to an exemplary embodiment, the present disclosure is directedto a method of collecting mononuclear cells, comprising providing anadsorption column through which whole blood is flowed to create purifiedwhole blood, and separating the purified whole blood into mononuclearcells and at least one component.

BRIEF DESCRIPTION OF THE DRAWINGS

Features, aspects, and advantages of the present embodiments will becomeapparent from the following description, appended claims, and theaccompanying exemplary embodiments shown in the drawings, which arebriefly described below.

FIG. 1 is a diagrammatic depiction of a separation system useful in theseparation and collection of mononuclear cells, according to anexemplary embodiment;

FIG. 2 is a perspective view of the front panel of a separation systemwith a disposable processing set for collecting mononuclear cellsmounted on the device, according to an exemplary embodiment;

FIG. 3 is a diagram showing the disposable processing set of FIG. 2,according to an exemplary embodiment;

FIG. 4 is a flow diagram illustrating an improved method for obtainingmononuclear cells, according to an exemplary embodiment;

FIG. 5 is a flow diagram illustrating an improved method for obtainingmononuclear cells, according to another exemplary embodiment; and

FIG. 6 is a flow diagram illustrating an improved method for obtainingmononuclear cells, according to another exemplary embodiment.

DETAILED DESCRIPTION

There are several aspects of the present subject matter which may beembodied separately or together in the devices and systems described andclaimed below. These aspects may be employed alone or in combinationwith other aspects of the subject matter described herein, and thedescription of these aspects together is not intended to preclude theuse of these aspects separately or the claiming of such aspectsseparately or in different combinations as set forth in the claimsappended hereto.

Some embodiments may provide for collecting MNCs with reduced plasmalipid interference during MNC harvest.

Some embodiments may provide for more accurate collection and harvest ofMNCs by allowing for a clearer interface between blood component layers.

During harvest of MNCs, non-target substances may be present in the MNCproduct that can interfere with accurate harvesting of the target MNCs.Plasma proteins and lipids may interfere, for example, in the event thatthe donor/patient has certain disease states or medications, such aselevated bilirubin levels and drugs such as mycophenolate mofetil (MMF)and cyclosporine, which cause hyperlipidemia.

FIG. 1 is a diagrammatic depiction of a separation system 10 useful inthe separation and collection of mononuclear cells, as described herein,and FIG. 2 shows an exemplary embodiment of the separation system 10,The system 10 may include a separation component 12 and a disposableprocessing kit 14 (FIG. 2) that is mounted thereon. Flow direction andrate may be controlled by a plurality of pumps 15 engaged with theprocessing kit 14, In one embodiment, the separation principle used bythe separator 12 is based on centrifugation, but an automated separatorbased on a different separation principle (e.g., spinning membrane,etc.) may also be used.

A patient may be connected to the fluid circuit 14, which may provide asterile closed pathway between the separation component 12 and theremainder of the processing kit 14, Whole blood that is withdrawn fromthe patient may be introduced into the separation component 12, wherethe whole blood may be separated to provide a target cell population,which in the context of the present disclosure may be mononuclear cells.Other components separated from the whole blood, such as red blood cellsand platelets may be returned to the patient or collected inpre-attached containers of the blood processing set. The separatedtarget cell population, e.g., mononuclear cells, may then be collectedfor future use or prepared for various therapies. One example of atherapy involving MNCs that may benefit from reducing plasma lipidinterference during MNC harvest is extracorporeal photopheresis or“ECP”. ECP involves the extracorporeal exposure of MNCs combined with aphotoactive compound, such as 8-methoxypsoralen or “8-MOP” which is thenphotoactivated by ultraviolet light, followed by re-infusion of thetreated MNCs. Removal of plasma lipids, which absorb UV light duringirradiation, may lead to generally more consistent and less variableirradiation procedures, thereby enhancing accuracy of irradiation dosingand shortening procedure time.

Apparatus useful in the collection of mononuclear cells, and providingthe separation component 12 of FIG. 1, include for example the Amicus®Separator made and sold by Fenwal, Inc., of Lake Zurich, Ill.Mononuclear cell collections using a device such as the Amicus® aredescribed in greater detail in U.S. Pat. No. 6,027,657, the contents ofwhich are incorporated by reference herein in its entirety. The fluidcircuit 14 (FIG. 3) may include a blood processing container 16 defininga separation chamber suitable for harvesting MNCs from whole blood.

As shown in FIG. 2, a disposable processing set or fluid circuit 14(which includes container 16) may be mounted on the front panel of theseparation component 12. The processing set (fluid circuit 14) mayinclude a plurality of processing fluid flow cassettes 23L, 23M and 23Rwith tubing loops for association with peristaltic pumps 15 on theseparation component 12. Fluid circuit 14 may also include a network oftubing and pre-connected containers for establishing flow communicationwith the patient and for processing and collecting fluids and blood andblood components, as shown in greater detail in FIG. 3.

As seen in FIG. 3, the disposable processing set 14 may include acontainer 60 for supplying anticoagulant, an in-process container 62, acontainer 64 for holding a crystalloid solution, such as saline, acontainer 66 for collecting plasma, and a container 68 for collectingthe mononuclear cells.

With reference to FIG. 3, fluid circuit 14 may include inlet line 72, ananticoagulant (AC) line 74 for delivering AC from container 60, an RBCline 76 for conveying red blood cells from chamber 16 of set 14 tocontainer 67, a platelet-poor plasma (PPP) line 78 for conveying PPP tocontainer 66 and line 80 for conveying mononuclear cells to and fromseparation chamber 16 and collection container 68.

The blood processing set may also include one or more venipunctureneedle(s) for accessing the circulatory system of the patient. As shownin FIG. 3, fluid circuit 14 may include inlet needle 70 and returnneedle 82. In an alternative embodiment, a single needle may serve asboth the inlet and outlet needle.

Fluid flow through fluid circuit 14 may be driven, controlled andadjusted by a microprocessor-based controller in cooperation with thevalves, pumps, weight scales and sensors of separation component 12 andfluid circuit 14, the details of which are described in the previouslymentioned U.S. Pat. No. 6,027,657.

A separation chamber may be defined by the walls of the processingcontainer 16. The processing container 16 may comprise two differentcompartments 16 a and 16 b (FIG. 3). Using both compartments 16 a and 16b for separation in a procedure may enable multiple target products tobe separated simultaneously and/or multiple steps to be completedsimultaneously. If only one compartment is used for separation, theother compartment may optionally be used as an in-process, waste, orstorage container. In operation, the separation device 12 may rotate theprocessing container 16 about an axis, creating a centrifugal fieldwithin the processing container 16. Details of the mechanism forrotating the processing container 16 are disclosed in U.S. Pat. No.5,360,542 titled “Centrifuge with Separable Bowl and Spool ElementsProviding Access to the Separation Chamber,” which is also incorporatedherein by reference in its entirety.

In one embodiment, an apheresis device or system 10 may include aprogrammable controller that is pre-programmed with one or moreselectable protocols. A user/operator may select a particular processingprotocol to achieve a desired outcome or objective. The pre-programmedselectable protocol(s) may be based on one or more fixed and/oradjustable parameters. During a particular processing procedure, thepre-programmed controller may operate the separator 12 and processingchamber 16 associated therewith to separate blood into its variouscomponents, as well as operate one or more pumps to move blood, bloodcomponents and/or solutions through the various openable valves andtubing segments of a processing set, such as processing set 14illustrated in FIG. 3. The various processing steps performed by thepre-programmed automated apheresis device may occur separately, inseries, simultaneously or any combination of these.

An automated apheresis device may be used to perform MNC collection in abatch process in which MNCs continuously collect in the chamber 16 untilthe target cycle volume is reached. During the continuous collection ofMNCs within the chamber 16, different blood components separate intolayers that may be detected by an optical interface detector thatmonitors the location and presence of the interface between layers.Details of an exemplary mechanism for interface detection are disclosedin U.S. Pat. No. 6,027,657, the contents of which are incorporated byreference herein in its entirety. Before and during the transfer of theMNCs out of the chamber 16, MNCs and other blood components (e.g.,plasma, etc.) may pass through an optical sensor 17, located downstreamof the chamber 16, which detects the presence of cells in the tubingline to determine the start and end of the MNC harvest (i.e. when toopen and close the valves leading to the product container). The term“downstream” describes an event proximal to post-separation, and theterm “upstream” describes an event proximal to pre-separation.“Downstream” and “upstream” are relative terms, with the reference pointbeing the time/location of separation. After MNC harvest is complete,the remaining cells in the line may be flushed into the productcontainer with a predetermined volume of plasma known as the “plasmaflush”.

The ability of the interface detector to accurately detect the interfacebetween blood component layers may be facilitated by removal ofnon-target substances (e.g., plasma proteins and lipids) that may bepresent in the blood that can interfere with the separation procedure.Additionally, the removal of non-target substances may improve theability of the optical sensor 17 to accurately detect the presence ofcells in the tubing line to determine the start and end of the MNCharvest to facilitate precise harvesting of the target MNCs.

EXAMPLES

Without limiting any of the foregoing, the subject matter describedherein may be found in one or more methods, systems and/or products. Forexample, in a first aspect of the present subject matter, an improvedsystem and method for obtaining MNCs is set forth in FIG. 4. The inletneedle 70 of FIG. 3 attached to inlet line 72 may first be connected toa blood source 5 (e.g, donor, patient, blood bag, etc.). Whole blood mayenter the separation chamber 16 of the separator 12, which separates thewhole blood into plasma and cellular components. The plasma may beseparated and directed into a plasma container 66, and the cellularcomponents may be separated and directed into a different container 67.The separated cellular contents may be combined with a replacement fluid69 that has minimal non-target content (e.g., plasma proteins and/orlipids) that may interfere with optical sensor readings. Examples ofsuitable replacement fluids include fresh frozen plasma, immunoglobulinsolution, albumin, and/or other colloid solutions. The cellularcomponents mixed with replacement fluid may then be returned to theseparation chamber 16, where target MNCs may be collected. The targetMNCs may be harvested into a designated container 68 to be processed forfurther treatment. Non-target components may be collected or returned tothe patient/donor.

The process and steps of whole blood initially entering the separationchamber 16 and the cellular components and replacement fluid mixreturning to the separation chamber 16 portrayed in FIG. 4 may takeplace substantially in series if only one compartment 16 a or 16 b isutilized. Alternatively, the process and steps of whole blood enteringthe separation chamber 16 and the fluid mix returning to the separationchamber 16 may take place substantially at the same time if bothcompartments 16 a and 16 b are utilized. In an embodiment in which theprocesses take place substantially in series, whole blood entering oneof the compartments 16 a or 16 b may separate into plasma and cellularcomponents, both of which may be directed to separate containers untilseparation of plasma and cellular components is complete. An opticalsensor 17 may optionally be placed downstream of the separation chamber16 a and/or 16 b at a tubing line leading to the MNC product container68 and/or leading to the plasma container 66 to determine when plasma isclear enough and plasma diversion can stop. Subsequently, the cellularcomponents may combine with the replacement fluid within one of thecompartments 16 a or 16 b, and further separation into target MNCs andnon-target components may take place.

In an embodiment in which the steps of whole blood entering theseparation chamber 16 and the fluid mix returning to the separationchamber 16 take place substantially at the same time, whole bloodentering a first compartment (e.g., 16 a) may separate into plasma andcellular components, with the plasma being sent to an plasma container62. Simultaneously, the cellular components may join the replacementfluid and together enter a second compartment (e.g., 16 b) and therefurther separate into target MNCs and non-target components. An opticalsensor 17 may optionally be placed downstream of the separation chamber16 a at a tubing line leading to the MNC product container 68 and/orleading to the plasma container 66 to determine when plasma is clearenough and plasma diversion can stop. As the replacement fluid continuesto enter compartment 16 b and the clarity of the plasma leavingcompartment 16 a improves sufficiently as determined by the opticalsensor 17, the plasma diversion from compartment 16 a into the plasmacontainer 66 can be stopped, and any unseparated whole blood, includingthe contents of compartment 16 a, may be directed to compartment 16 b tocontinue MNC collection.

In another aspect of the present subject matter, an improved method forobtaining MNCs is set forth in FIG. 5. An inlet needle may first beconnected to a blood source 5 (e.g, donor, patient, blood bag, etc.) atstep 100 of FIG. 5. In step 200, whole blood enters a separator, whichseparates the whole blood into plasma (step 300 a) and cellularcomponents (step 300 b). In the embodiment in FIG. 5, the separator ofstep 200 may be a centrifugal or spinning membrane separator. Anexemplary spinning membrane and hardware is disclosed in greater detailin PCT Patent Application No. PCT/US2012/28492, which is incorporatedherein by reference in its entirety, although any suitable membraneassembly may be used, Plasma separated in step 300 a may be flowedthrough an adsorption column in step 400. An exemplary adsorption columnis the MONET filter made and sold by Fresenius Medical Care. Anotherexemplary adsorption column is disclosed in greater detail inInternational Publication No, WO 2012/141697 and U.S. Pat. No.6,569,112, each of which is hereby incorporated by reference herein inits entirety, although any suitable column may be used. In step 500,purified plasma from the adsorption column in step 400 may be combinedwith the cellular components of step 300 b. The combined product of step500 may then be separated in a separation chamber of a separator in step600 to collect and harvest MNC target cells separated from non-targetcomponents.

In another aspect of the present subject matter, an improved method forobtaining MNCs is set forth in FIG. 6. An inlet needle may first beconnected to a blood source 5 (e.g, donor, patient, blood bag, etc.) atstep 101 of FIG. 6. In step 201, whole blood enters a whole bloodadsorption column, which removes certain lipids, proteins, antibodies,and/or fatty acids in step 301. An exemplary whole blood adsorptioncolumn is the DALI® adsorber made and sold by Fresenius Medical Care,although any suitable whole blood adsorption column may be used.Purified whole blood exiting from the WB adsorption column of step 201may then be separated in a separation chamber 16 of a separator in step401 to collect and harvest MNC target cells separated from non-targetcomponents.

The embodiments disclosed herein are for the purpose of providing adescription of the present subject matter, and it is understood that thesubject matter may be embodied in various other forms and combinationsnot shown in detail. Therefore, specific embodiments and featuresdisclosed herein are not to be interpreted as limiting the subjectmatter as defined in the accompanying claims.

1-11. (canceled)
 12. A method of collecting mononuclear cells,comprising: separating whole blood into plasma and cellular components;purifying the plasma through a plasma adsorption column to createpurified plasma; combining the cellular components with the purifiedplasma to form a first mixture; and separating the first mixture intomononuclear cells and at least one component.
 13. The method of claim12, further comprising the step of performing extracorporealphotopheresis on the mononuclear cells, and wherein the purified plasmacontains fewer proteins, lipids, and/or bilirubin than does the plasma.14. The method of claim 12, wherein the step of separating whole bloodinto plasma and cellular components is performed by a centrifugalseparator.
 15. The method of claim 12, wherein step of separating wholeblood into plasma and cellular components is performed by a spinningmembrane separator.
 16. The method of claim 12, wherein the mononuclearcells comprise at least one of lymphocytes, monocytes, and stem cells.17. A method of collecting mononuclear cells, comprising: providing anadsorption column through which whole blood is flowed to create purifiedwhole blood; and separating the purified whole blood into mononuclearcells and at least one component.
 18. The method of claim 17, whereinthe purified whole blood is separated by a centrifugal separator. 19.The method of claim 17, wherein the mononuclear cells comprise at leastone of lymphocytes, monocytes, and stem cells.
 20. The method of claim17, wherein the purified whole blood contains fewer proteins, lipids,and/or bilirubin than does the whole blood.
 21. A blood separationsystem comprising: a processing kit including an adsorption column, afirst separation chamber, and a second separation chamber; and aseparation component including a pump system and a controller, whereinthe controller is configured to control the pump system to convey wholeblood into the first separation chamber to separate the whole blood intoplasma and cellular components, convey the plasma through the adsorptioncolumn to create purified plasma, combine the cellular components withthe purified plasma to form a first mixture, and convey the firstmixture into the second separation chamber to separate the first mixtureinto mononuclear cells and at least one component.
 22. The bloodseparation system of claim 21, further comprising a photopheresis deviceconfigured to perform extracorporeal photopheresis on the mononuclearcells, wherein the purified plasma contains fewer proteins, lipids,and/or bilirubin than does the plasma.
 23. The blood separation systemof claim 21, wherein the first separation chamber is defined by acentrifugal separation container.
 24. The blood separation system ofclaim 21, wherein the first separation chamber is defined by a spinningmembrane separator.
 25. The blood separation system of claim 21, whereinthe mononuclear cells comprise at least one of lymphocytes, monocytes,and stem cells.
 26. A blood separation system comprising: a processingkit including an adsorption column and a blood processing container; anda separation component including a pump system and a controller, whereinthe controller is configured to control the pump system to convey wholeblood through the adsorption column to create purified whole blood, andconvey the purified whole blood into the blood processing container toseparate the purified whole blood into mononuclear cells and at leastone component.
 27. The blood separation system of claim 26, wherein theblood processing container comprises a centrifugal separation container.28. The blood separation system of claim 26, wherein the mononuclearcells comprise at least one of lymphocytes, monocytes, and stem cells.29. The blood separation system of claim 26, wherein the purified wholeblood contains fewer proteins, lipids, and/or bilirubin than does thewhole blood.